Proposal Summary (Abstract) Anti-CD19 chimeric antigen receptor (CAR) T cells work well in treating patients with acute lymphoblastic leukemia (ALL), but much less so in other more indolent B cell malignancies, such as chronic lymphocytic leukemia (CLL). Improved CAR T cells are necessary to enhance antitumor efficacy in the treatment of these cancers. Our lab has recently demonstrated that constitutive expression of the co-stimulatory molecule CD40 ligand (CD40L, CD154) improves cytotoxicity and antitumor efficacy of CAR T cells in vitro and in an immune- compromised xenograft mouse model. My hypotheses are that CD40L-modified CAR T cells promote an improved antitumor response through several mechanisms: (i) by enhancing the activation of CAR T cells resulting in improved cytotoxicity and functional CAR T cell persistence; (ii) by increasing the immunogenicity of tumor cells leading to their recognition by host immune effector cells; and/or (iii) via activation of host immune cells to mediate a sustained endogenous antitumor response. The long-term objectives of this proposed study aim to investigate the biology of CD40L-modified CAR T cells to provide mechanistic insights into how CD40L-modified CAR T cells achieve an enhanced antitumor efficacy when compared to T cells expressing the CAR alone. The first specific aim will be to transfer the CD40L-modified CAR T cell platform into a fully immune-competent syngeneic lymphoma mouse model and to determine its antitumor efficacy. Based on preliminary data, we anticipate that the results of these proposed experiments will validate the use of CD40L-modified CAR in this setting. The second specific aim will determine the effect CD40L-modified T cells have on the immunogenic phenotype of tumor cells and their clearance. The third specific aim will be to investigate the ability of CD40L-modified CAR T cells to change the cellular composition of the tumor microenvironment and to induce an antitumor response by endogenous T cells. The implications of this work will extend beyond its application in indolent B cell malignancies, but also provide insight into how genetic engineering of T cells can alter the tumor microenvironment and provide a rationale for utilizing this CD40L- based CAR T cell approach in solid tumors.